A new analytical method is developed and appropriately validated by means of a high performance liquid chromatography (HPLC) in reverse phase to quantify pantoprazole in pharmaceutical preparations. The HPLC determination was performed on a reversed phase column (Inertsil ODS 53m; 150 x 4.6 mm i.d) using a mobile phase (0.6 mL min-1 ) with UV-detection at 289 nm. A rectilinear relationship between mean peak area and concentration of PNT was observed in the range 25-200 μg mL-1 with a detection limit of 6.0 μg mL-1 and a quantitation limit of 20.0 μg mL-1. Intra-day and interday precision, and accuracy of the methods have been established according to the current ICH guidelines. The results were statistically compared with those of the reference method by applying Student’s t-test and F-test. Accuracy, evaluated by means of the spike recovery method was in the range 97.7-102.2%, with precision (RSD) better than 2%.